The overall aim of this SCCOR application is to understand the complex events related to ventilator-associated lung injury (VALI). In Project 6, we will determine the role of oxidative stress in VALI. Oxidative stress has been generally implicated in acute lung injury through production of reactive oxygen species (ROS). To detoxify ROS, lung cells express a network of integrated antioxidant enzymes (AOE) consisting of both classical and phase II detoxifying enzymes. However, if the oxidant stress is overwhelming, ROS may suppress or inactivate the AOE system resulting in cellular damage. We have recently shown by genetic and functional analysis that NF-E2 related factor 2 (Nrf2) plays a critical role in hyperoxia-induced lung injury. This transcription factor is involved in lung epithelial cell repair and stretch and regulates the induction of phase II AOEs. We have also shown that Nrf2-knockout mice are more susceptible to hyperoxia-induced lung injury. In collaboration with the various Cores of the SCCOR, genomic analysis revealed an upregulation of Nrf2 and AP-1 family members, such as fos-related protein-1 (Fra-1), as well as specific AOEs in the lungs of mice and dogs exposed to mechanical ventilation (MV). Thus, we hypothesize that activation of transcription factors such as Nrf2 and Fra-1 and their downstream effectors plays a critical role in the modulation of VALI. To test this hypothesis, we propose the following Specific Aims: 1) Determine the molecular responses of the lung in response to MV and hyperoxia in the development of VALI. We will compare gene and protein expression profiles and markers of injury in the lungs of mice and dogs exposed to MV and hyperoxia; 2) Examine the role of Nrf2 and identify its downstream effectors in VALI; and 3) Examine the role of Fra-1 and identify its downstream effectors in VALI. For these studies, we will use respective knockout and/or transgenic mice with specific lung overexpression of mutant or wildtype proteins of Nrf2 and Fra-1, as well as genomics and proteomics techniques in collaboration with the various Cores. Cross species comparison and validation of these results in human will be done through collaboration with other Basic and Clinical Research Projects, respectively. Results of these studies will provide further insight into the role of oxidative stress as well as the molecular and genetic basis of VALI. The identification of molecular targets will aid in the development of new strategies aimed at minimizing the potential I hazardous effects of MV and hyperoxia.